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The fungus Aspergillus niger is among the most abundant fungi in the world and is widely used as a cell factory for protein and metabolite production. This fungus forms asexual spores called conidia that are used for dispersal. Notably, part of the spores and germlings aggregate in an aqueous environment. The aggregated conidia/ germlings give rise to large microcolonies, while the nonaggregated spores/germlings result in small microcolonies. Here, it is shown that small microcolonies release a larger variety and quantity of secreted proteins compared to large microcolonies. Yet, the secretome of large microcolonies has complementary cellulase activity with that of the small microcolonies. Also, large microcolonies are more resistant to heat and oxidative stress compared to small microcolonies, which is partly explained by the presence of nongerminated spores in the core of the large microcolonies. Together, it is proposed that heterogeneity in germination and aggregation has evolved to form a population of different sized A. niger microcolonies, thereby increasing stress survival and producing a meta-secretome more optimally suited to degrade complex substrates.

Solid-state NMR (ssNMR) spectroscopy facilitates the non-destructive characterization of structurally heterogeneous biomolecules in their native setting, for example, comprising proteins, lipids and polysaccharides. Here we demonstrate the utility of high and ultra-high field 1 H-detected fast MAS ssNMR spectroscopy, which exhibits increased sensitivity and spectral resolution, to further elucidate the atomic-level composition and structural arrangement of the cell wall of Schizophyllum commune, a mushroom-forming fungus from the Basidiomycota phylum. These advancements allowed us to reveal that Cu(II) ions and the antifungal peptide Cathelicidin-2 mainly bind to cell wall proteins at low concentrations while glucans are targeted at high metal ion concentrations. In addition, our data suggest the presence of polysaccharides containing N-acetyl galactosamine (GalNAc) and proteins, including the hydrophobin proteins SC3, shedding more light on the molecular make-up of cells wall as well as the positioning of the polypeptide layer. Obtaining such information may be of critical relevance for future research into fungi in material science and biomedical contexts.

Germination of conidia is an essential process within the Aspergillus life cycle and plays a major role during the infection of hosts. Conidia are able to avoid detection by the majority of leukocytes when dormant. Germination can cause severe health problems, specifically in immunocompromised people. Aspergillosis is most often caused by Aspergillus fumigatus (A. fumigatus) and affects neutropenic patients, as well as people with cystic fibrosis (CF). These patients are often unable to effectively detect and clear the conidia or hyphae and can develop chronic non-invasive and/or invasive infections or allergic inflammatory responses. Current treatments with (tri)azoles can be very effective to combat a variety of fungal infections. However, resistance against current azoles has emerged and has been increasing since 1998. As a consequence, patients infected with resistant A. fumigatus have a reported mortality rate of 88% to 100%. Especially with the growing number of patients that harbor azole-resistant Aspergilli, novel antifungals could provide an alternative. Aspergilloses differ in defining characteristics, but germination of conidia is one of the few common denominators. By specifically targeting conidial germination with novel antifungals, early intervention might be possible. In this review, we propose several morphotypes to disrupt conidial germination, as well as potential targets. Hopefully, new antifungals against such targets could contribute to disturbing the ability of Aspergilli to germinate and grow, resulting in a decreased fungal burden on patients.

Biofilm formation during infections with the opportunistic pathogen Aspergillus fumigatus can be very problematic in clinical settings, since it provides the fungal cells with a protective environment. Resistance against drug treatments, immune recognition as well as adaptation to the host environment allows fungal survival in the host. The exact molecular mechanisms behind most processes in the formation of biofilms are unclear. In general, the formation of biofilms can be categorized roughly in a few stages; adhesion, conidial germination and development of hyphae, biofilm maturation and cell dispersion. Fungi in biofilms can adapt to the in-host environment. These adaptations can occur on a level of phenotypic plasticity via gene regulation. However, also more substantial genetic changes of the genome can result in increased resistance and adaptation in the host, enhancing the survival chances of fungi in biofilms. Most research has focused on the development of biofilms. However, to tackle developing microbial resistance and adaptation in biofilms, more insight in mechanisms behind genetic adaptations is required to predict which defense mechanisms can be expected. This can be helpful in the development of novel and more targeted antifungal treatments to combat fungal infections.

Abstract: Therapeutic options to treat invasive fungal infections are still limited. This makes the development of novel antifungal agents highly desirable. Naturally occurring antifungal peptides represent valid candidates, since they are not harmful for human cells and are endowed with a wide range of activities and their mechanism of action is different from that of conventional antifungal drugs. Here, we characterized for the first time the antifungal properties of novel peptides identified in human apolipoprotein B. ApoB-derived peptides, here named r(P)ApoBLPro, r(P)ApoBLAla and r(P)ApoBSPro, were found to have significant fungicidal activity towards Candida albicans (C. albicans) cells. Peptides were also found to be able to slow down metabolic activity of Aspergillus niger (A. niger) spores. In addition, experiments were carried out to clarify the mechanism of fungicidal activity of ApoB-derived peptides. Peptides immediately interacted with C. albicans cell surfaces, as indicated by fluorescence live cell imaging analyses, and induced severe membrane damage, as indicated by propidium iodide uptake induced upon treatment of C. albicans cells with ApoB-derived peptides. ApoB-derived peptides were also tested on A. niger swollen spores, initial hyphae and branched mycelium. The effects of peptides were found to be more severe on swollen spores and initial hyphae compared to mycelium. Fluorescence live cell imaging analyses confirmed peptide internalization into swollen spores with a consequent accumulation into hyphae. Altogether, these findings open interesting perspectives to the application of ApoB-derived peptides as effective antifungal agents. Key points: Human cryptides identified in ApoB are effective antifungal agents.ApoB-derived cryptides exert fungicidal effects towards C. albicans cells.ApoB-derived cryptides affect different stages of growth of A. niger. Graphical abstract[Figur

Malassezia yeasts are lipid dependent and part of the human and animal skin microbiome. However, they are also associated with a variety of dermatological conditions and even cause systemic infections. How these yeasts can live as commensals on the skin and switch to a pathogenic stage has long been a matter of debate. Lipids are important cellular molecules, and understanding the lipid metabolism and composition of Malassezia species is crucial to comprehending their biology and host-microbe interaction. Here, we investigated the lipid composition of Malassezia strains grown to the stationary phase in a complex Dixon medium broth. In this study, we perform a lipidomic analysis of a subset of species; in addition, we conducted a gene prediction analysis for the detection of lipid metabolic proteins. We identified 18 lipid classes and 428 lipidic compounds. The most commonly found lipids were triglycerides (TAG), sterol (CH), diglycerides (DG), fatty acids (FAs), phosphatidylcholine (PC), phosphatidylethanolamine (PE), ceramides, cholesteryl ester (CE), sphingomyelin (SM), acylcarnitine, and lysophospholipids. Particularly, we found a low content of CEs in Malassezia furfur, atypical M. furfur, and Malassezia pachydermatis and undetectable traces of these components in Malassezia globosa, Malassezia restricta, and Malassezia sympodialis. Remarkably, uncommon lipids in yeast, like diacylglyceryltrimethylhomoserine and FA esters of hydroxyl FAs, were found in a variable concentration in these Malassezia species. The latter are bioactive lipids recently reported to have antidiabetic and anti-inflammatory properties. The results obtained can be used to discriminate different Malassezia species and offer a new overview of the lipid composition of these yeasts. We could confirm the presence and the absence of certain lipid-biosynthesis genes in specific species. Further analyses are necessary to continue disclosing the complex lipidome of Malassezia species and the impa

Malassezia is a lipid-dependent genus of yeasts known for being an important part of the skin mycobiota. These yeasts have been associated with the development of skin disorders and cataloged as a causal agent of systemic infections under specific conditions, making them opportunistic pathogens. Little is known about the host-microbe interactions of Malassezia spp., and unraveling this implies the implementation of infection models. In this mini review, we present different models that have been implemented in fungal infections studies with greater attention to Malassezia spp. infections. These models range from in vitro (cell cultures and ex vivo tissue), to in vivo (murine models, rabbits, guinea pigs, insects, nematodes, and amoebas). We additionally highlight the alternative models that reduce the use of mammals as model organisms, which have been gaining importance in the study of fungal host-microbe interactions. This is due to the fact that these systems have been shown to have reliable results, which correlate with those obtained from mammalian models. Examples of alternative models are Caenorhabditis elegans, Drosophila melanogaster, Tenebrio molitor, and Galleria mellonella. These are invertebrates that have been implemented in the study of Malassezia spp. infections in order to identify differences in virulence between Malassezia species.